Conformational changes probably mediate both macromolecular recognition and the conversion of chemical free energy into work during protein biosynthesis, but the putative structural changes have not been confirmed or characterized. Our investigation will focus on the recognition and binding of aminoacyl-tRNA by the ribosome, since this process requires precise recognition, utilizes energy, and is likely to involve conformational changes. Functional fluorescent-labeled analogs of aminoacyl-tRNA will be used to characterize spectrally the structural and functional states of aminoacyl-tRNA in both ribosomal and aa-tRNA. EF-tu.GTP complexes, and to obtain information on the topology of the complexes. The kinetics of aminoacyl-tRNA binding to ribosomes and movement from site to site will be determined by monitoring changes in fluorescence as a function of time in order to examine, among other things, both the catalytic effect of elongation factor Tu and the effect of GTP hydrolysis on the process. The distance moved by a tRNA in a GTP-hydrolysis-dependent shift from one site to another on the ribosome will be measured directly by fluorescence energy transfer between two fluorescent-labeled tRNAs. When this data is combined with other distance measurements, the molecular movement which is coupled to an exergonic chemical reaction can be quantitated. The details of these processes will provide clues to the molecular mechanisms which control the general processes of macromolecular recognition and free energy conversion in cells, and the reasons why those mechanisms occasionally fail.